VL 31 (Ralph Gräf) Flashcards
Connecting the nuclear lamina with the outside world
LINC complex
* SUN domain: inner nuclear envelope membrane
* KASH domains: outer nuclear envelope membrane
Association of LINC complexes with all cytoskeletal elements
→Linkage with the nuclear envelope
* interaction: cell contacts - cytoskeletal elements
* mech. connection of nuclei of neighboring cells
Desmosome composition:
Proteoglycans build proteoglycan aggregates:
Structure
* hundreds of glycan side chains bound to core protein (~ 2000 aa)
* ca. 100 proteoglycan side chains bound to one core glycan
Proteoglycans:
* form solid gel
* H2O-binding
* formation of a hydrated matrix for fibrous proteins
* structural variability
Multiadhesive components: Laminin
- basal lamina component
- important in cell migration (→ neuron development)
- heterotrimer; 3 different SU
- MW: 800.000
- Different binding domains for ECM components
Multiadhesive components: Fibronectin
- RGD = Arg-Gly-Asp; Integrin-binding site
- In connective tissue, blood, basal lamina
- Important for cell migration, adhesion, wound healing
- Composed of 3 types of repetitive aa sequences
- 2 SU → homodimer; MW: 550 kDa
- Various binding domains for ECM-components
Nuclear composition
- double membrane; outer membrane connected to ER
- nuclear pores = large protein complexes allowing passage of large molecules through nuclear envelope
- nuclear lamina beneath inner membrane consists of lamins (= IF proteins)
- chromatin with DNA
- nucleolus (ribosome biogenesis)
Terms around DNA and chromatin
-
Chromosome:
DNA molecule(s) + associated proteins; prior to S-
phase 1 molecule, after S-phase 2 DNA molecules -
Chromatid:
1 DNA molecule + associated proteins -
Chromatin:
total nuclear DNA together with packaging proteins -
Euchromatin:
transcriptionally active, almost uncondensed
chromatin (genes) -
Heterochromatin:
transcriptionally inactive, condensed chromatin -
Centromer:
connecting site of sister chromatids consisting of
condensed DNA with specific associated proteins. -
Kinetochore:
centromeric substructure maturating only during
mitosis (before prekinetochore). Defines the attachment site for spindle microtubules. -
Centrosome:
microtubule-organizing center, nucleation and
binding site for spindle microtubules. -
Telomere:
specialized structure at each chromosome end that protects ends and ensures complete replication of chromosome ends during S-phase
Nucleolus
- Non-membranous complex; with proteins & RNA
- 1-5/cell
- Site of
–> rDNA transcription
–> pre-rRNA processing o ribosome SU assembly - ribosomal SU assembly in GC → ribonucleoproteins
- decomposition prior to mitosis → de novo formation at NORs (nucleolus organizing regions) at rRNA-gene loci
- FCs of the later nucleoli arise at NORs
Chromosome structure:
Histones and epigenetics:
- epigenetic regulation: stable, inheritable regulation of chromosomal functions through information not directly encoded by DNA
- posttranslational histone modifications → chromatin (non)-accessible
Histones and epigenetics – heterochromatin formation:
- e.g. X-chromosome inactivation in women (barr body formation)
- binding of methyl cytosine-binding protein MeCP2 at CpG-islands→recruits histone methyltransferase
- H3K9me (tri-methylation)→HP1-binding
- HP1 recruits histone-, DNA-modifying enzymes (H3K9 histone methyltransferase) promoting heterochromatin
formation, transcription repression - result: men, women with same X-chromosomal transcription level
Further chromosome-associated proteins
- non-histone proteins: discovered after DNA-, histone-extraction from metaphase chromosomes
- →remaining material forms chromosome scaffold composed of non-histone proteins
Picture
* same structure of SMC1/3
–> alpha-helical structure
–> hinge domain → collapsing coiled-coil structure
–> head domain interaction with kleisins (Scc1)
* Scc1: ATPase
–> ATP cleavage→ring opening
–> ATP bound→ring closure
- Strong ring model: DNA strands held together after S-phase by ring junction→favored
- Weak ring model:
–> Handcuff model (left): each DNA strand with cohesin ring; linkage via additional protein
–> Chain link model (right)
Centromeres & kinetochores:
- CENP-A: H3 isoform within a centromere-specific nucleosome (ubiquitous in eukaryotes)
- CENP-T/W/S/X form a further centromere-specific nucleosome
- Both = adaptor for Ndc80 complex
- motor proteins (kinesin, dynein) at kinetochore
- Kinetochore regulation through mitotic checkpoint apparatus
- > 65 proteins (many “CENP” proteins)
Organization of the nuclear envelope:
Lamins: structure and assembly
- Simple building block: lamin dimer
- Lateral dimer assembly (head-tail) → tetramer in assembly
region; antiparallel assembly → no polarity - Hexamer in region of assembled dimers; tetramer in length
Nuclear Lamina
- Synthese von Proproteinen im Cytoplasma = Prälamin A, B1, B2
- Prälamin mit NLS
- Bindung von NLS von Importin-alpha
- Assemblierung der Lamine nach Ablösen von Importin-alpha
- Caax-Box am C-Terminus (C = Cys, aa = 2 aliphatische AS-Reste, X = irgendwelche AS)
- CaaX-Box bei kleinen GTPasen und anderen Proteinen→Isoprenylierung, nicht bei Lamin C
- Iosprenreste: Farnesyl, Geranyl-Geranyl
➔ Modifikationen von Cys durch Farnesyltransferase (FTase) am ER - Proteolytische Spaltung von aaX durch RCE1 (rass-converting enzyme) oder Zmpste24, redundante Enzyme
- Methylierung durch ICMT (Isoprenyl-Cystein-Methyl-Transferase)
- Proteolytische Spaltung der letzten 15 AS bei Prälamin A durch Zmpfste24→keine sterische Hinderung
- Isoprenylrest→Bindung an Kernhüllenmembran
- A-Typ-Laminnetzwerke unterhalb der B-Typ-Lamine
- Interaktion mit Transmembranproteinen der inneren Kernhüllenmembran
